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WO2005062914A2 - Dispositif d'echantillonnage microbien - Google Patents

Dispositif d'echantillonnage microbien Download PDF

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Publication number
WO2005062914A2
WO2005062914A2 PCT/US2004/043254 US2004043254W WO2005062914A2 WO 2005062914 A2 WO2005062914 A2 WO 2005062914A2 US 2004043254 W US2004043254 W US 2004043254W WO 2005062914 A2 WO2005062914 A2 WO 2005062914A2
Authority
WO
WIPO (PCT)
Prior art keywords
collection fluid
collection
integrated
fluid
microbial
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/US2004/043254
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English (en)
Other versions
WO2005062914A3 (fr
Inventor
Mansour Samadpour
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Institute for Environmental Health Inc
Original Assignee
Institute for Environmental Health Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Institute for Environmental Health Inc filed Critical Institute for Environmental Health Inc
Priority to US10/584,098 priority Critical patent/US9957475B2/en
Publication of WO2005062914A2 publication Critical patent/WO2005062914A2/fr
Publication of WO2005062914A3 publication Critical patent/WO2005062914A3/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/30—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration
    • C12M41/36—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration of biomass, e.g. colony counters or by turbidity measurements
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M37/00—Means for sterilizing, maintaining sterile conditions or avoiding chemical or biological contamination
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/02—Devices for withdrawing samples
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/02—Devices for withdrawing samples
    • G01N2001/028—Sampling from a surface, swabbing, vaporising
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/38—Diluting, dispersing or mixing samples
    • G01N2001/383—Diluting, dispersing or mixing samples collecting and diluting in a flow of liquid

Definitions

  • Preferred aspects of the present invention relate generally to collecting samples of microbial agents, and particularly to improving the efficiency of collecting representative and reproducible samples of microbial agents on surfaces, and from air.
  • Sponge sampling requires execution of a prolonged series of procedural steps to ensure that the sampling media: is aseptically extracted from its packaging prior to use; is applied to the surface in a way that collects microbial contamination in a representative manner; and is returned to the packaging in such a way as to prevent cross-contamination and loss of sample integrity.
  • Variations in technique such as pressure during application, number of 'passes,' and/or whether or not an 'area template' is used, can result in an undesirable level of variable results.
  • excision sampling requires execution of a prolonged series of procedural steps to ensure that the instruments used to excise the sample are sterile, and that a portion of the sample is excised in a way that collects microbial contamination in a representative manner, and that the sample is packaged in such a way as to prevent cross-contamination and loss of sample integrity.
  • removal of a portion of the surface by excision can be detrimental to the aesthetic quality and desirability of the food product.
  • variations in the depth and technique of the excision can lead to a greater or lesser mass of food product being included in the sample, thereby making it difficult to interpret and normalize the data in terms of microbial contamination per unit surface area.
  • Preferred aspects of the present invention provide a device and method, which can be used for air sampling as well as for surface sampling. Additional preferred aspects of the present invention provide a universal sampling device and methods for using same, allowing repeated aseptic sampling of microbial agents on surfaces and from air. Particularly preferred aspects provide for integrated sanitization of the sampling unit between sampling runs or events. Specific aspects provide improved devices and methods for conducting microbial sampling of surfaces including food surfaces, food contact surfaces, and non-food contact surfaces. Particular aspects provide a device which reduces the number of procedural steps required to conduct microbial sampling of surfaces. Additional preferred aspects provide a device that collects microbial contamination with a high efficiency. Further aspects provide a device and method, which reduces the variation in sample collection, thereby reducing the variation and error in the results.
  • Yet further aspects provide a device which yields results that can be correlated with historical data. Particularly preferred aspects provide a device which does not degrade the aesthetic quality and desirability of the surface being sampled. Additional aspects provide a device which can be used for collection of single discrete samples. In preferred aspects, the inventive devices 'self-sanitize' for repeated sampling. Particularly preferred aspects provide a device which facilitates the collection of a composite sample, representing the sum of several single discrete samples.
  • an appropriate quantity of the fluid is delivered and dispersed onto the surface to be sampled (e.g., by spray delivery through a nozzle).
  • the fluid entrains any microbial contamination that may be present, and is then recovered by application of a vacuum to the targeted surface, drawing the fluid from the surface and directing it to a collection reservoir.
  • the contents of the reservoir are emptied after a single sample, or, optionally after multiple samples have been collected in forming one composite sample.
  • a nozzle delivering the microbial sampling fluid is incorporated into a sampling head.
  • the sampling head may have a well-defined geometry, such as a rectangular configuration, which facilitates/complements the collection of a sample from a known surface area.
  • Specific preferred embodiments of the present invention provide an apparatus for sampling microbial organisms present on surfaces, comprising: a reservoir suitable for providing microbial collection fluid; a sterilizable sample collection chamber; a sterilizable, integrated collection fluid delivery and collection fluid recovery member, suitable to deliver collection fluid to a target surface, and contemporaneously recover the delivered fluid from the surface; delivery means, in communication with both the reservoir and the integrated member, and operable to aseptically deliver collection fluid from the reservoir to the integrated member; and vacuum means, in communication with both the sample collection chamber and the integrated member, and operable to direct collection fluid, delivered and recovered by the integrated member, to the sample collection chamber.
  • the integrated fluid delivery and recovery member is reversibly detachable.
  • the reservoir is a pressurizable chamber.
  • the delivery means comprises a compressor in communication with the chamber.
  • the delivery means comprises a fluid pump.
  • the vacuum means comprises a vacuum pump, and a moisture trap interposed between the sample collection chamber and the vacuum pump.
  • the integrated collection fluid delivery and collection fluid recovery member comprises a spray nozzle suitable to direct sample collection fluid toward the target surface.
  • the integrated collection fluid delivery and collection fluid recovery member comprises a actuatable valve for actuated delivery of the sample collection fluid.
  • the sampling apparatus further comprises a sanitizing means for sanitizing the integrated collection fluid delivery and collection fluid recovery member.
  • the sanitation means comprises a sanitation unit having a sanitizing reservoir for receiving the integrated collection fluid delivery and collection fluid recovery member.
  • the integrated collection fluid delivery and collection fluid recovery member conforms to the target surface contour.
  • the target surface is a food surface or a food-contact surface.
  • the food surface is that of an animal or animal carcass.
  • the animal carcass is bovine, porcine, equine or avian.
  • the microbial collection fluid preserves microbial vitality without promoting microbial growth, allowing for determination of microbial number per unit surface area.
  • the microbial collection fluid promotes microbial growth, allowing for determination of a presence of absence of surface microbial organisms.
  • Particularly preferred aspects of the present invention comprise combining the instant inventive surface sampling methods with the advanced pathogen testing and carcass-certification methods for slaughter operations described in WO04098296A2, which is incorporated by reference herein in its entirety.
  • FIG. 1 shows a schematic diagram of a preferred embodiment of the invention, illustrating the use of a compressor to pressurize peptone water, a microbial sampling fluid, for delivery through a nozzle contained in a sampling wand/head. The sampling fluid is collected in a sample bottle by means of applying a vacuum.
  • Figure 2 shows a schematic diagram of an exemplary sanitizing unit used to prepare (e.g., sanitize) a sampling wand before and between collection of samples.
  • Figure 3 shows a schematic diagram of a sampling pattern used during comparative experiments disclosed herein.
  • Figure 4 shows a trend-line comparison of results of sampling of meat surfaces between traditional swab sampling, and sampling according to preferred aspects of the present invention.
  • Figure 5 shows a trend-line comparison between the results of sampling of a
  • an exemplary inventive sampling device 1 comprises a vacuum pump 2, a compressor 4, a pressurized tank 6 containing microbial sampling fluid, a vacuum application/suction head 8 containing a nozzle 10 for the application of the microbial sampling fluid, a vacuum hose 12 for returning the applied sampling fluid to a sample bottle 14, and a moisture trap 16 for protecting the vacuum pump 2.
  • the compressor 4 and pressurized tank 6 could be replaced, for example, with a non-pressurized tank in communication with a pump operable to deliver sampling fluid from the tank to the nozzle 10 of the application/suction head 8.
  • the principle operation of this preferred embodiment of the invention is to direct (e.g., spray) microbial sampling fluid onto the surface to be sampled, and then recover the liquid into a sample bottle 14 by means of, for example, application of a vacuum at the application/suction head 8.
  • Preferred microbial sampling solutions include those which preserve the viability of the organism without promoting growth. Samples based on such solutions can be returned to the laboratory and analyzed by standard plating techniques to determine the number of viable organisms per unit of recovered sampling solution.
  • microbial sampling solutions By measuring the total volume of recovered sampling solution, the total number of organisms recovered can be computed. By measuring the total surface area where the sampling device was applied, the number of viable organisms per unit of area can be computed.
  • Preferred microbial sampling solutions also include those which preserve the viability of the organism while promoting growth (so called "enrichment broths"). Samples based on these solutions can be returned to the laboratory and analyzed to determine the presence/absence of organisms of concern.
  • Appropriate microbial sampling solutions include, but are not limited to, those described standard liquid media such as those found in the media index of the "Bacteriological Analytical Manual" published by the U.S.
  • the compressor 4 provides pressure to the pressure tank 6 that delivers the microbial sampling fluid to the spray nozzle 10 located inside the application/suction head 8 (integrated collection fluid delivery and collection fluid recovery member 8).
  • the spray nozzle 10 directs the fluid onto the surface to be sampled, in a line approximately the width of the application/suction head 8.
  • the rate of application which along with the time of application affects the volume of fluid recovered per unit surface area, can be adjusted by modifying the pressure in the pressure tank 6.
  • a pump can be placed between the reservoir of the microbial sampling fluid and the application nozzle 10, and the rate of application can be adjusted by adjustment of the pump rate.
  • the fluid is extracted from the surface being sampled by applying a vacuum through a flexible hose 12 terminating in a vacuum head (e.g., application/suction head 8) containing the spray nozzle 10.
  • the microbial sampling fluid is extracted from the surface being sampled by vacuum, first through a wand (e.g., application/suction head 8), and then a hose 12 connecting the wand to the sample bottle.
  • the wand handle 18 connecting the wand 8 and the sample hose 12 to the sample bottle 14 has, for example, a trigger design valve that controls delivery and/or amount of fluid delivered to the target surface.
  • An optional screening means e.g., stainless steel screen 24
  • An optional moisture trap 16 between the sample bottle 14 and the vacuum pump 2 protects the vacuum pump from moisture.
  • the shape of the integrated collection fluid delivery and collection fluid recovery member 8 facilitates collection of a sample from a known sample area.
  • all components are contained on a cart or other movable platform.
  • the sampling bottle 14 and moisture trap 16 are preferably located in a accessible rack on top of the cart. Optimally, the rack has space for additional sample bottles.
  • the vacuum pump 2 , compressor 4, and pressure vessel 6 are preferably located on the enclosed bottom shelf. In particular embodiments, a double-door provides access to the equipment on the shelf.
  • the moisture trap 16, sample bottle 14, sample hose 12, and wand 8 can quickly be separated.
  • the various segments of the sampling solution supply line 20 supplying peptone water from the pressure tank 6 to the spray nozzle 10 in the suction head 8 are optionally connected by 'quick disconnects' 22.
  • an optionally retractable electric cord that is located inside the covered portion of the cart supplies power to the unit.
  • the vacuum pump 2 is optionally controlled by a switch on the cart side panel.
  • a separate sanitizing unit 30 is constructed to allow the wand 8 to be sterilized before and/or between sampling events.
  • the apparatus of the sanitizing unit 30 is best understood by reference to FIGURE 2.
  • the sanitizing unit 30 contains a sanitizing reservoir 32 of liquid (e.g., water) maintained at an elevated temperature (e.g., by a heater 36) sufficient to be lethal to microorganisms upon contact.
  • liquid e.g., water
  • the wand 8 with the vacuum tube 12 (or a detachable portion thereof) is removed from the sampling device 1, and each end is connected to the sanitizing unit 30 in such a fashion as to form a closed loop between the wand 8, the heated water reservoir 32 and a sanitizer pump 34.
  • Activation of the pump 34 causes hot liquid (e.g., water) to be circulated through the wand 8, ensuring that all microbial organisms therein contained are killed.
  • the wand 8 is returned to the sampling device 1, along with a fresh sterilized sampling bottle 14, and the sampling unit 1 is ready to collect another sample.
  • a diffuser tube is connected to the receiving hose in the collection chamber 14, which will convert the sampling device to an 'impinger,' allowing for quantitative air sampling.
  • the collection chamber is partially filled with microbial sampling fluid.
  • Impingers are well known in the art. Typically, they comprise special glass tubes designed to collect airborne contaminants by bubbling the sampled air at a high flow rate through a method specific absorbing liquid. The liquid used can then be analyzed to determine airborne contaminate levels.
  • liquid-filled impingers have been preferred as devices for collecting bioaerosols.
  • Liquid-filled impingers offer the advantage of easy microbial analysis because portions of the collection liquid can be placed directly onto nutrient media (agar), which is incubated and the resulting colonies enumerated and identified. Impingers are widely available, inexpensive, and sterilizable.
  • EXAMPLE 1 The exemplary apparatus 1 of FIGURE 1 was compared to current art swab sampling methods for collection of a microbial sample from a meat surface. Meat surfaces were freshly prepared by thinly slicing a whole top round portion of beef using a sterilized knife. The internal tissue of such whole rounds is generally regarded as having a low microbial organism level.
  • FIGURE 2 The results were analyzed by conducting a 'rend-line' comparison, as shown in FIGURE 4. An excellent correlation was observed, with a slope of near unity. Additionally, the inventive apparatus and method showed less variation when sampling this rough surface.
  • EXAMPLE 2 The apparatus 1 of FIGURE 1 was compared to current art-recognized swab sampling for collection of a microbial sample from a hard surface. A FORMICATM surface was inoculated at two different levels (10 5 and 10 4 CFU/100 cm 2 ) with Biotype I Escherichia coli
  • ECC ECC
  • slurry of sterilized fecal matter prepared from known stock laboratory cultures in slurry of sterilized fecal matter. Surfaces were inoculated by spreading 10 mL of the slurry on a 300 cm 2 area of the surface to be sampled, according to the schematic provided in FIGURE 3.
  • Swab samples were collected using sterile sponges moistened with 25 mL of 0.1% peptone water, according to standard methodology recommended by the United States Department of Agriculture (USDA), Food Safety and Inspection Services (FSIS).
  • USDA United States Department of Agriculture
  • FSIS Food Safety and Inspection Services
  • the apparatus 1 of FIGURE 1 was used to collect comparative samples.
  • the ECC content of the collected samples was determined through standard serial dilution and plate counting methods. The results obtained with each

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Abstract

Certains aspects de la présente invention concernent un nouvel appareil et de nouveaux procédés permettant d'échantillonner des organismes microbiens présents sur des surfaces. L'appareil préféré de cette invention comprend: un réservoir pouvant fournir un fluide de prélèvement microbien; une chambre de prélèvement d'échantillons pouvant être stérilisée; un élément de distribution de fluide de prélèvement et de récupération de fluide de prélèvement intégré et pouvant être stérilisé, lequel élément peut distribuer un fluide de prélèvement sur une surface cible et récupérer simultanément le fluide distribué à partir de la surface; une unité de distribution communiquant à la fois avec le réservoir et l'élément intégré et pouvant distribuer de manière aseptique le fluide de prélèvement du réservoir vers l'élément intégré; et une unité de vide communiquant à la fois avec la chambre de prélèvement d'échantillons et l'élément intégré et pouvant orienter le fluide de prélèvement, distribué et récupéré par l'élément intégré, vers la chambre de prélèvement d'échantillons. Des aspects supplémentaires de cette invention concernent un échantillonnage rapide et à haut débit d'organismes microbiens présents sur des surfaces et consistant: à distribuer le fluide de prélèvement d'échantillons sur une surface cible et à récupérer simultanément le fluide distribué de la surface cible au moyen d'un élément intégré de distribution de fluide de prélèvement et de récupération de fluide de prélèvement; et à prélever le fluide de prélèvement d'échantillons récupéré dans une chambre de prélèvement d'échantillons communiquant avec l'élément intégré, le prélèvement d'échantillons étant au moins partiellement réalisé. Les procédés et l'appareil de cette invention peuvent être appliqués tant à l'échantillonnage de surface qu'à l'échantillonnage atmosphérique.
PCT/US2004/043254 2003-12-22 2004-12-22 Dispositif d'echantillonnage microbien Ceased WO2005062914A2 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
US10/584,098 US9957475B2 (en) 2003-12-22 2004-12-22 Microbial sampling device

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US53136403P 2003-12-22 2003-12-22
US60/531,364 2003-12-22

Publications (2)

Publication Number Publication Date
WO2005062914A2 true WO2005062914A2 (fr) 2005-07-14
WO2005062914A3 WO2005062914A3 (fr) 2005-11-17

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Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2011006170A2 (fr) * 2009-07-10 2011-01-13 Microbial-Vac Systems, Inc. Système et procédé de nettoyage d'un équipement de recueil d'agent contaminant

Family Cites Families (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4038057A (en) 1974-05-13 1977-07-26 Andersen 2000, Inc. Closed circuit sampler
US6013227A (en) * 1997-12-17 2000-01-11 Johnson & Johnson Medical, Inc. Lumen device reprocessor without occlusion
FR2779822B1 (fr) * 1998-06-10 2000-09-08 Millipore Sa Procede et appareil de prelevement pour l'analyse microbiologique de l'air
US6338282B1 (en) * 1998-06-19 2002-01-15 Agro-Enviro Consultants, Inc. Portable liquid sampling system
US6472203B1 (en) 1999-11-01 2002-10-29 Environmental Microbiology Laboratory, Inc. Combination air sampling cassette and nutrient media dish
US7100461B2 (en) * 2002-02-27 2006-09-05 Microbial-Vac Systems, Inc. Portable contaminant sampling system
US7487689B2 (en) * 2003-01-15 2009-02-10 Tracetrack Technology Ltd. Contaminant scanning system
US7919232B2 (en) 2003-05-05 2011-04-05 Institute For Environmental Health, Inc. Advanced pathogen testing and carcass-certification methods for slaughter operations

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Publication number Publication date
WO2005062914A3 (fr) 2005-11-17
US9957475B2 (en) 2018-05-01
US20080166752A1 (en) 2008-07-10

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